Follicular fluid (FF) includes various biologically active proteins which can affect follicular growth and maturation. Certain proteins could reflect the physiological and pathological status of follicles. The aim of the present study was to explore the key proteins associated with pathogenesis of follicular cysts, some of which may be candidate biomarkers for the condition. We analysed the proteomes of FF from small, medium, large and cystic follicles by two-dimensional electrophoresis (2-DE) combined with mass spectrometry (MS). The protein components in FF were found to be significantly different among groups; about 300 proteins spots in each group were examined, and 32 differentially expressed proteins were identified from different groups. To further reveal the source of identified proteins, transcripts encoding two of these, transferrin and RBP-4, were detected in granulosa cells (GCs) by RT-PCR, as well as the proteins were detected in 24 h culture media of GCs by ELISA. High levels of RBP-4 were examined in FF of cystic follicles by 2-DE analysis, which were significantly different to those in large follicles (p < 0.05). In conclusion, the study enriches our understanding of the proteins of FF; RBP-4 and transferrin originate from passive transfer and follicular synthesized secretion, and RBP-4 might be a candidate biomarker for porcine follicular cysts. Combined with histological studies, these results further suggest that changes of the type and quantity of proteins in FF might be attributed to an abnormal metabolism of follicular cells and structure of follicular wall in cystic follicles. Our findings will contribute to further insight into the pathogenesis of follicular cysts.

To investigate the causes of the occurrence and persistence of porcine cystic follicles, we evaluated the apoptosis and proliferation of follicular cells in these cysts. Apoptotic frequencies were examined by TUNEL assay and the expression of apoptosis regulators (XIAP, bax, bc1-2 and caspase-3) by immunohistochemistry, Western blotting and real-time quantitative PCR; cell proliferation activity was evaluated by PCNA immunohistochemistry and proliferation of in vitro cultured granulosa and theca cells. The low apoptotic frequency and weak proliferative activity were found in cystic follicles. Low frequency of apoptosis might be associated with decreased amounts of apoptotic-related factors (bax and caspase-3) and increased amounts of anti-apoptotic factors (XIAP and bcl-2) in cystic follicles. Significantly lower proliferation activity was detected in granulosa and theca cells from cystic follicles, and lesser PCNA-positive cells were found in cystic follicles. Our results indicate that the programmed cell death and cell proliferation system were altered in cystic follicles. The disorder between apoptosis and proliferation was responsible for maintaining a static condition without degeneration, which leads to the long-term persistence of follicles. These findings provide important novel insights into the pathogenesis of follicular cysts in sows.